Characterisation of Ca2+ membrane permeability of renal A6 cells upon different osmotic conditions.
The nature of the calcium-transporting mechanisms involved in A6 cell calcium homeostasis under iso- and hypo-osmotic conditions was investigated using fura-2 (AM) as a cell calcium indicator. Under steady-state conditions, intracellular calcium (Ca2+i) was increased by Bay K8644 or by gramicidin, an ionophore which depolarises A6 cell membranes. The Ca2+i increase following calcium addition (to calcium-depleted cells) or membrane depolarisation was blocked by nifedipine but not by verapamil or omega-conotoxin, indicating that the membrane calcium permeability may be mediated by voltage-dependent and dihydropyridine-sensitive calcium channels. Ca2+i could also be increased by a hypo-osmotic shock having a linear relationship with the osmolarity change. This osmotically induced Ca2+i increase had an extracellular origin since it was absent when cells were suspended in a calcium-free medium and it was not affected by thapsigargin or TMB-8 application. In addition, it was inhibited by the calcium channel inhibitor, nifedipine. Furthermore, under hypo-osmotic conditions, an additional Ca2+i increase, sensitive to nifedipine, was measured when cells were depolarised by gramicidin or K-gluconate addition. It is proposed that the hypo-osmotically induced cell calcium increase implies the activation of voltage-dependent and nifedipine-sensitive calcium channels, presenting the same pharmacological characteristics as those involved in cell calcium homeostasis under iso-osmotic conditions. The initial Ca2+i increase was transient and stabilised to a value nevertheless higher than the iso-osmotic level; this secondary and incomplete regulatory phase did not occur in the presence of thapsigargin or TMB-8, thus providing evidence of intracellular calcium storage.
Brochiero
E
E
Department of Cellular and Molecular Biology, CEA-URA 1855 (CNRS), Laboratoire Jean-Maetz, Villefranche/Mer, France.
Ehrenfeld
J
J
D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester / Q000494:pharmacology; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002120:Calcium Channel Agonists / Q000494:pharmacology; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism*; D002463:Cell Membrane Permeability / Q000187:drug effects / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005707:Gallic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D006096:Gramicidin / Q000494:pharmacology; D007424:Intracellular Fluid / Q000187:drug effects / Q000378:metabolism; D017136:Ion Transport; D007686:Kidney Tubules, Distal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008564:Membrane Potentials / Q000187:drug effects; D009994:Osmolar Concentration; D019284:Thapsigargin / Q000494:pharmacology; D014982:Xenopus laevis
D000900:Anti-Bacterial Agents; D002120:Calcium Channel Agonists; D002121:Calcium Channel Blockers; D015220:Calcium Channels; D004791:Enzyme Inhibitors; D006096:Gramicidin; C006014:8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate; D005707:Gallic Acid; D019284:Thapsigargin; D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; D002118:Calcium